PROJECT SUMMARY The cytochrome P450s (CYPs) are the major enzymes involved in drug metabolism and bioactivation. It is well known that several CYP enzymes metabolize omega-3 fatty acids to their epoxy metabolites that inhibit angiogenesis, tumor growth, and metastasis. Numerous polycyclic aromatic hydrocarbons (PAH) are human carcinogens. PAH-DNA adducts may lead to DNA damage and mutations in critical genes, eventually leading to cancer. A significant positive linear regression between levels of PAH-DNA adducts and tumor incidence was observed in animal experiments in our laboratory. We also discovered that omega-3 fatty acids eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) inhibited CYP1B1, EZH2, DNMT3a, miR 17, miR19b-1 and significantly decreased pulmonary and hepatic PAH-DNA adducts, and tumor incidence. The central hypothesis of this application is that omega 3-fatty acids and their epoxy metabolites will attenuate pulmonary carcinogenesis by multiple mechanisms entailing attenuation of PAH-DNA adduct formation by modulating CYPs, as well as by suppression of tumorigenesis, probably via modulation of epigenetic genes (e.g., EZH2, DNMT3a, miR-17, miR-19b-1). We propose the following Specific Aims. Aim 1: To test the hypothesis that CYP1A1 and CYP1B1 play mechanistic roles in prevention of PAH carcinogenesis in mice maintained on EPA, DHA, or EPA + DHA diets, compared to those on a CO diet, followed by exposure of these mice to BP for the study of the mechanisms. Aim 2: To test the hypothesis that mice deficient in soluble epoxide hydrolase (sEH) will confer more protection than WT mice to EPA/DHA-mediated prevention of PAH carcinogenesis, as sEH is known to rapidly hydrolyze epoxy metabolites such as 17,18-epoxy eicosatetraenoic acid (EEQ) and 19,20-epoxy docosapentaenoic acids (EDP) in serum and tissues to inactive metabolites. In some experiments, we will treat WT mice with the specific sEH inhibitor, t-TUCB, or a new t-TUCB-like inhibitor (that is likely to go to human clinical trials soon), followed by treatment of mice with EPA/DHA and BP. Aim 3: To test the hypothesis that endogenous omega-3 fatty acids, especially their epoxy metabolites, will play a pivotal role in the prevention of pulmonary carcinogenesis by PAHs in vivo, and that there is a mechanistic link between CYP1, and sEH. Fat-1-transgenic (Fat-1-Tg) mice, which will convert endogenous omega-6 fatty acids (rich in CO) into omega-3 fatty acids and decrease the ratios of omega-6/omega-3, will be used in this study. We will also create Fat-1-Tg/sEH-null mice for exploring the mechanisms by which CYP1 and sEH enzymes contribute to omega-3 fatty acid-mediated prevention of PAH carcinogenesis. If our hypothesis that CYP1 and sEH enzymes play important roles in omega-3 fatty acids, i.e. EPA/DHA-mediated prevention of PAH-induced cancers turns out to be correct, then it will break new grounds in the current understanding of human cancer prevention. If successful, the proposed studies should lead to novel mechanisms in dietary interventions against lung cancers induced by PAHs.